The multiprotein complex gamma-secretase proteolytically cleaves the intramembrane region of amyloid precursor protein (APP), which in turn forms the plaques found in Alzheimer's disease (AD) patients. The catalytic component of gamma-secretase is the intramembrane aspartyl protease (IAP) called presenilin. Mutations in presenilin are directly linked to familial early-onset AD. Another known member of the IAP family is signal peptide peptidase (SPP), which functions to further proteolyze remnant signal peptides after they have been cleaved by signal peptidase. Knowledge of the biochemistry and function of individual SPPs are only beginning to be elucidated, and homologues are found in all kingdoms of life. Presenilin and SPP exhibit significant sequence similarity, strongly suggesting they share structural and catalytic features. Thus, a molecular understanding of the more tractable SPP will likely impact drug design for presenilin and gamma- secretase. The goal of this proposal is to express, characterize, and solve the crystal structure of an extremophilic bacterial SPP ortholog by itself, with a transition-state analog inhibitor and with a substrate mimic. In addition, drug candidates will be screened in silico. This first structure of an intramembrane protease will provide critical insight into the biochemistry of intramembrane proteolysis and enable structure- based AD drug development and screening.